The immune reconstitution inflammatory syndrome (IRIS) is a frequent complication of antiretroviral therapy in HIV+ individuals. IRIS is described as an inflammatory reaction at the site of infection with numerous opportunistic pathogens, caused by a recovery of the immune system. Although IRIS appears to be immune-mediated, the pathogenesis of this syndrome has not been studied in detail. Progressive Multifocal Leukoencephalopathy (PML) occurring in the context of IRIS is characterized by contrast enhancement, swelling and even mass effect that may prove lethal and may require treatment with steroids. While JC virus, the causative agent of PML, infects oligodendrocytes and astrocytes in the CNS white matter, newly recognized JCV-infected cerebellar granule cell neurons may also act as a target of the immune response in IRIS. Furthermore, we have identified intracortical lesions of PML and, for the first time, observed JCV infection of cerebral hemispheric cortical neurons. As anti-HIV medications become more potent, PML IRIS is likely to become more prominent clinically. We have defined the cellular immune response to JCV mediated by CD8+ T lymphocytes. While CD4+ T cell-mediated immune recovery appears to be of importance in IRIS, whether these cells react specifically against JCV has not been characterized in detail. In addition, the prevalence of JCV infection of neurons, and whether these cells can be recognized by the immune system and participate to IRIS, remains unknown. We hypothesize that rather than the absolute rise of CD4+ T cells during HAART, the frequency of JCV-specific CD4+ T cells will predict the development of IRIS in PML patients. PML lesions will have a different metabolic profile in IRIS cases by proton MR spectroscopy (1H-MRS) and reversible inflammation will be differentiated from permanent demyelination using magnetization transfer, diffusion and perfusion imaging. We predict that intracortical lesions of PML are an important part of the disease burden, including in some cases JCV-infected cortical neurons. Absence of MHC class I antigens on JCV-infected neurons may promote escape from immune recognition in IRIS. To test these hypotheses, we propose to perform the following experiments: 1) Analyze JCV-specific CD4+ T cells by Enzyme-linked immuno-spot (ELISPOT) and intracellular cytokine staining (ICS) assays in PML patients with and without IRIS 2) Evaluate the metabolism and differentiate inflammation from demyelination in PML lesions and adjacent gray matter of patients with an without IRIS with contrast-enhanced MRI, 1H-MRS, Magnetization Transfer, Diffusion and Perfusion Imaging 3) Explore whether JCV-infected neurons in cerebral cortex and granule cell layer of the cerebellum can be recognized by the immune system, determine their prevalence and characterize the molecular determinants of JCV tropism for these cells PUBLIC HEALTH RELEVANCE: JC virus (JCV) causes a demyelinating disease of the brain in immunosuppressed individuals called Progressive Multifocal Leukoencephalopathy (PML). In these studies, we will characterize the mechanisms leading to the potentially lethal immune reconstitution inflammatory syndrome (IRIS), which occurs frequently in HIV+ PML patients after initiation of antiretroviral therapy. The knowledge we will gain from these studies will greatly expand our understanding of JCV pathogenesis and will have a direct impact on patient care.